Immune evasion by Trypanosoma cruzi, the causative agent of Chagas disease is likely multifactorial. However, one key parasite derived factor implicated in immune evasion is cruzain (or cruzipain), the major protease of this organism. Our laboratory has successfully targeted this protease with a cysteine protease inhibitor. We can block the parasite life cycle in experimental animals treated with this inhibitor. We have also isolated a cruzain deficient parasite clone via drug selection. This clone, Cai-KR is non-pathogenic in wildtype mice, but can cause acute disease in RAG1-/- mice. Our preliminary results suggest that cruzain could play a role in preventing macrophage activation by degrading NF-kB in the cytoplasm of infected cells. In this application, we will use molecular, biochemical and cellular approaches to determine if T. cruzi interferes with NF-kB signaling and describe its mechanism. We will also compare the expression profile of T. cruzi infected macrophages with macrophages classically and alternatively activated with cytokines, as well as infected with intracellular bacteria. Finally, we propose to make a transgenic mouse over expressing cruzain to determine its effects on the immune system in vivo.